An understanding of the molecular basis of action, or mechanism, of proteolytic enzymes is a long-term objective of biochemical investigation, realization of which would have multiple benefits with regard to medical problems associated with protein metabolism. For metalloproteases, new substrates are being developed to aid in assay for proteolytic activity. Novel inhibitors and inactivators are being designed as probes of mechanism, and as potential leads to biomedically efficacious agents. Independent developments in structural biochemistry (crystallography) have created fresh opportunities by identifying new subcategories of metalloproteases, specifically dual-metal ion aminopeptidases and astacin- type proteases, for which kinetic investigations being undertaken are greatly needed for clarification of mechanism. Materials developed in this project should have applicability to a range of current challenges in protein chemistry, including hormone processing, collagen degradation, and invasive cancer.